Not applicable.
Not applicable.
Not Applicable.
(1) Field of the Invention
The present invention provides an aqueous composition comprising a Pseudomonas species, an alkali metal nitrate, an ethoxylate nonionic surfactant, and optionally monoammonium phosphate as a buffer in an aqueous solution. The Pseudomonas sp. is stable during storage of the composition and the composition is useful for cleaning or bioremediation applications.
(2) Description of Related Art
The use of Pseudomonads for bioremediation of biodegradable materials is very well known. Illustrative of the patent art are U.S. Pat. No. 3,963,576 to Horsfall et al.; U.S. Pat. No. 4,391,887 to Baumgarten et al.; U.S. Pat. No. 4,508,824 to Olsen; U.S. Pat. No. 4,672,037 to Daggett; U.S. Pat. No. 4,673,505 to Wong; U.S. Pat. No. 4,822,490 to Dyadechko et al.; U.S. Pat. No. 4,910,143 to Vandenbergh; U.S. Pat. No. 4,970,000 to Eppler et al. and U.S. Pat. No. 5,494,580 to Baskys et al.
However, because the pseudomonad compositions are dried, such as by lyophilization, or frozen with preservatives, there is a significant die-off of the bacteria over time (usually about 75% by cell count) due to the preservation. Further, the cultures generally need to be reactivated in a bulk starter medium in order to increase the number and viability of the cells. The use of a bulk starter media for this purpose is well known to those skilled in the art. To circumvent the problem of die-off and to avoid the need for a bulk starter media, U.S. Pat. No. 5,980,747 to Vandenbergh discloses an aqueous pseudomonad composition which is storage-stable at room temperatures for storage, shipment, and use and yet immediately active without the use of a bulk starter.
In bioremediating a site or cleaning a surface, the microorganism culture suspension is applied to the site or surface in situ or in the case of some bioremediation activities, above ground. Commonly, in bioremediation, material from the site is combined with the cultured microorganism in a reaction vessel. Surfactants are usually added to the reaction vessel to reduce the surface tension of the contents of the reaction which facilitates contact between the material to be degraded and the microorganisms. Surfactants are also commonly used in bioremediation of a site or cleaning a surface. Thus, the cleaning ability of the microorganism-containing solution is principally attributed to the surfactant content in the reaction vessel. In general, microorganisms do not tolerate surfactants well. Therefore, for most bioremediation or cleaning applications, the microorganisms are stored separately from the surfactants (See U.S. Pat. No. 4,349,633 to Worne et al. for example).
However, it would be more convenient to provide a single composition that contains both the microorganisms and the surfactant. U.S. Pat. No. 6,387,874 to Schalitz et al. discloses a composition consisting of spores from the genus Bacillus in a mixture containing surfactants. The Bacillus spores have a long shelf life and protect the Bacillus from inactivation by surfactants. U.S. Pat. No. 6,248,234 to Cline provides a solid dissolvable bioactive element containing both surfactants and microorganisms.
In light of the above, there remains a need for an aqueous pseudomonad composition for cleaning or bioremediation which includes a surfactant and which is stable over long periods of time for storage, shipment, and use and yet immediately active without the use of a bulk starter. It would be further desirable that the composition be inexpensive to produce and store.
The present invention provides a composition useful for cleaning and bioremediation applications comprising a Pseudomonas species, an alkali metal nitrate, an ethoxylate nonionic surfactant, and optionally monoammonium phosphate as a buffer in an aqueous solution. The Pseudomonas sp. is stable during storage of the composition.
In particular, the present invention provides an aqueous composition containing a live bacterium of the genus Pseudomonas useful for degrading biological material comprising (a) water; (b) at least one species of the Pseudomonas at between about 106 to 1010 CFU per mL of the water; (c) an ethoxylate nonionic surfactant; and (d) an alkali metal nitrate in an amount between about 0.1% and 5% by weight of the water and optionally a buffering amount of monoammonium phosphate so that the Pseudomonas species is preserved in a living form at 25xc2x0 C.
In a preferred embodiment of the composition, the Pseudomonas species is a Pseudomonas putida and in particular, the Pseudomonas putida selected from the group consisting of Pseudomonas putida NRRL-B-18118, Pseudomonas putida NRRL-B-15078, and mixtures thereof. In particular embodiments, the composition further includes a Pseudomonas fluorescens, preferably, the Pseudomonas fluorescens deposited as NRRL-B-21658.
In a preferred embodiment of the composition, the alkali metal is sodium and when the monoammonium phosphate is included, it is included in a weight ratio between 1 to 1 and 125 to 1 alkali metal nitrate to monoammonium phosphate.
In further embodiment of the composition, the ethoxylate nonionic surfactant is a linear alcohol ethoxylate. Preferably, the ethoxylate nonionic surfactant is a (C9-C11) alkyl alcohol ethoxylate, CAS number 68439-46-3. It is preferable that the ethoxylate nonionic surfactant is at a concentration of between about 0.1% and 1%.
The present invention also provides a method for degradation of a biological material comprising (a) providing an aqueous composition which includes (1) water; (2) at least one species of the Pseudomonas at between about 106 to 1010 CFU per mL of the water; (3) an ethoxylate nonionic surfactant; and (4) an alkali metal nitrate in an amount between about 0.1% and 5% by weight of the water and optionally a buffering amount of monoammonium phosphate so that the Pseudomonas species is preserved in a living form at 25xc2x0 C.; (b) holding the composition at a temperature between about 8 and 35xc2x0 C. to provide the preserved Pseudomonas species; (c) inoculating the preserved Pseudomonas species into the biodegradable material; and (d) degrading the biological material with the Pseudomonas species.
In a preferred embodiment of the method, the Pseudomonas species is a Pseudomonas putida and in particular, the Pseudomonas putida selected from the group consisting of Pseudomonas putida NRRL-B-18118, Pseudomonas putida NRRL-B-15078, and mixtures thereof. In particular embodiments, the method further includes a Pseudomonas fluorescens, preferably, the Pseudomonas fluorescens deposited as NRRL-B-21658.
In a preferred embodiment of the method, the alkali metal is sodium and when the monoammonium phosphate is included, it is included in a weight ratio between 1 to 1 and 125 to 1 alkali metal nitrate to monoammonium phosphate.
In further embodiment of the method, the ethoxylate nonionic surfactant is a linear alcohol ethoxylate. Preferably, the ethoxylate nonionic surfactant is a (C9-C11) alkyl alcohol ethoxylate, CAS number 68439-46-3. It is preferable that the ethoxylate nonionic surfactant is at a concentration of between about 0.1% and 1%.
Therefore, it is an object of the present invention to provide an aqueous pseudomonad composition which is stable over long periods of time at room temperatures and in the presence of surfactants and which is immediately effective for cleaning or bioremediation applications.
It is a further object of the present invention to provide an aqueous pseudomonad composition that is inexpensive to prepare.
These and other objects of the present invention will become increasingly apparent with reference to the preferred embodiments.